Project Summary/Abstract The urea cycle is the major pathway for detoxification of ammonia in mammals. Arginase 1 deficiency is thought to be the least common of the urea cycle disorders and results in hyperargininemia. In humans, deficiency of this enzyme is characterized clinically by progressive mental impairment, spasticity, and growth retardation, with only periodic episodes of hyperammonemia unlike the other urea cycle disorders where this is much more common. In recent experiments, the Lipshutz Lab has found substantial anatomical, ultrastructural and electrophysiological differences between knockout animals and wild type controls. Constitutive and global arginase deficiency led to decreased intrinsic excitability, altered functional synaptic transmission, decreased dendritic arborization and decreased synapse density. Some of these measures (but not all) were rescued by hepatic gene therapy that controlled plasma arginine. In addition, and unexpectedly, heterozygotes showed intermediate neuronal findings where plasma biochemistry is not different than arginase wildtype mice, suggesting an intrinsic neuronal or cell autonomous role for neuronal arginase activity. These measurable differences at the neuron, synapse, and circuit level have begun to elucidate the functional abnormalities in arginase deficiency. This proposal will examine the hypothesis that arginase expression in the central nervous system (CNS) plays an important role in the developing CNS. In addition to mapping arginase expressing neurons in the brain, selective arginase 1 loss in neurons will be induced to assess whether this leads to unique functional deficits. Preliminary data: The Lipshutz lab and collaborators have (amongst other findings): 1) constructed and characterized the arginase 1 knockout mouse; 2) demonstrated long-term survival and rescue with recombinant adeno-associated viral vectors; 3) demonstrated that only low-level ureagenesis is necessary for long-term survival; 4) shown that peripheral metabolism can result in control of circulating plasma arginine; and 5) shown that loss of arginase gene expression results in abnormalities of intrinsic excitability and the development of the dendritic arbor of neurons. In Aim 1, mapping of arginase-expressing neurons in the brain will be performed using the tissue-clearing technology of CLARITY. In Aim 2, the hypothesis that neuron-specific loss of arginase recapitulates the anatomical, electrophysiological, and behavioral abnormalities seen in the constitutive knockout mouse will be tested. Successful completion of the proposed studies will provide a greater molecular understanding of and the mechanism behind the alterations in the brain, neurons, and synapses in arginase 1 deficiency and hyperargininemia.